Department of Neurosurgery, Section Experimental Neurosurgery/Tumor Immunology, University Hospital Carl Gustav Carus, TU Dresden, Fetscherstr. 74, 01307, Dresden, Germany.
Institute for Clinical Genetics, Faculty of Medicine Carl Gustav Carus, TU Dresden, Fetscherstr. 74, 01307, Dresden, Germany.
BMC Cancer. 2017 Dec 28;17(1):889. doi: 10.1186/s12885-017-3932-y.
Survivin, belonging to the inhibitor of apoptosis (IAP) gene family, is abundantly expressed in tumors. It has been hypothesized that Survivin facilitates carcinogenesis by inhibition of apoptosis resulting in improved survival of tumorigenic progeny. Additionally, Survivin plays an essential role during mitosis. Together with its molecular partners Aurora B, Borealin and inner centromere protein it secures bipolar chromosome segregation. However, whether increased Survivin levels contribute to progression of tumors by inducing chromosomal instability remains unclear.
We overexpressed Survivin in U251-MG, SVGp12, U87-MG, HCT116 and p53-deficient U87-MG and HCT116 cells. The resulting phenotype was investigated by FACS-assisted cell cycle analysis, Western Blot analysis, confocal laser scan microscopy, proliferation assays, spectral karyotyping and in a U251-MG xenograft model using immune-deficient mice.
Overexpression of Survivin affected cells with knockdown of p53, cells harboring mutant p53 and SV40 large T antigen, respectively, resulting in the increase of cell fractions harboring 4n and >4n DNA contents. Increased γH2AX levels, indicative of DNA damage were monitored in all Survivin-transduced cell lines, but only in p53 wild type cells this was accompanied by an attenuated S-phase entry and activation of p21. Overexpression of Survivin caused a DNA damage response characterized by increased appearance pDNA-PKcs foci in cell nuclei and elevated levels of pATM S1981 and pCHK2 T68. Additionally, evolving structural chromosomal aberrations in U251-MG cells transduced with Survivin indicated a DNA-repair by non-homologous end joining recombination. Subcutaneous transplantation of U251-MG cells overexpressing Survivin and mycN instead of mycN oncogene alone generated tumors with shortened latency and decreased apoptosis. Subsequent SKY-analysis of Survivin/mycN-tumors revealed an increase in structural chromosomal aberrations in cells when compared to mycN-tumors.
Our data suggest that increased Survivin levels promote adaptive evolution of tumors through combining induction of genetic heterogeneity with inhibition of apoptosis.
Survivin 属于凋亡抑制因子（IAP）基因家族，在肿瘤中大量表达。有假说认为 Survivin 通过抑制凋亡从而促进肿瘤发生，使肿瘤前体的存活率提高，进而促进肿瘤的发生发展。此外，Survivin 在有丝分裂过程中发挥着重要作用。它与分子伴侣 Aurora B、Borealin 和内着丝粒蛋白共同确保了染色体的两极分离。然而，Survivin 水平的增加是否通过诱导染色体不稳定而促进肿瘤的进展仍不清楚。
我们在 U251-MG、SVGp12、U87-MG、HCT116 和 p53 缺失的 U87-MG 和 HCT116 细胞中转染 Survivin 以过表达 Survivin。通过流式细胞术辅助的细胞周期分析、Western Blot 分析、共聚焦激光扫描显微镜、增殖实验、光谱核型分析以及在免疫缺陷小鼠的 U251-MG 异种移植模型中研究所得表型。
Survivin 的过表达分别影响了 p53 敲低的细胞、携带突变型 p53 的细胞和 SV40 大 T 抗原的细胞，导致含有 4n 和 >4n DNA 含量的细胞分数增加。所有 Survivin 转导的细胞系中都监测到了增加的 γH2AX 水平，这表明存在 DNA 损伤，但只有在 p53 野生型细胞中，才会伴随着 S 期进入的减弱和 p21 的激活。Survivin 的过表达导致 DNA 损伤反应，表现为细胞核中 pDNA-PKcs 焦点的增加和 pATM S1981 和 pCHK2 T68 的水平升高。此外，U251-MG 细胞中转染 Survivin 后出现的结构性染色体畸变表明，非同源末端连接重组参与了 DNA 修复。过表达 Survivin 和 mycN 而不是单独过表达 mycN 癌基因的 U251-MG 细胞的皮下移植生成了潜伏期缩短和凋亡减少的肿瘤。随后对 Survivin/mycN 肿瘤进行 SKY 分析显示，与 mycN 肿瘤相比，细胞中的结构性染色体畸变增加。
我们的数据表明，Survivin 水平的增加通过诱导遗传异质性与抑制凋亡相结合，促进了肿瘤的适应性进化。
